Keenan is best known as the lead singer of the multi-platinum rock bands Tool and A Perfect Circle with whom he has released four and three studio albums, respectively. In 2003 he created Puscifer as a side project, financing and releasing its first studio album in October 2007. Since rising to fame, Keenan has been a noted recluse, although he does emerge to support charitable causes.

In addition to his music career, he has performed improvisational stand-up comedy and ventured into acting. He is the current owner of Merkin Vineyards and the associated winery, Caduceus Cellars, and has part ownership of Stronghold Vineyards, all located in Arizona, where he lives. Among other business ventures, he is a partner in the Los Angeles restaurant Cobras & Matadors, and owns a produce market and an organic market in Cornville, Arizona, the latter featuring a tasting room for his wines as well as a food court.

James Keenan was born in Ravenna, Ohio, on April 17, 1964, the only child of Judith Marie and Michael Loren Keenan. When his parents divorced in 1968, his father, a high school teacher, moved to Michigan and Keenan would only see him about once a year for the next 12 years. His mother remarried, bringing Keenan into an "intolerant and unworldly household", where his intelligence and creative expression would be stifled. When Keenan was 11, his mother suffered a paralyzing cerebral aneurysm, which would later serve as the inspiration for many of his creative works. A few years later, she persuaded Keenan to live with his father in Michigan. Keenan considers this "the best move [he] ever made."

Inspired by Bill Murray's performance in the 1981 comedy film Stripes, Keenan joined the United States Army, with the intention of having the G.I. Bill fund his dream of attending art school. By this point, he had lived in Ohio, Michigan, New Jersey, New York, Oklahoma, Kansas, and Texas. He initially served in the Army as a forward observer before studying at the United States Military Academy Preparatory School (West Point Prep School) from 1983 to 1984. In addition to completing a rigorous math and English curriculum, he ran on the cross country team and sang in the glee club. It was during his time in the military that he adopted the sobriquet "Maynard" on a whim. He was distinguished in basic and advanced training, but declined an appointment to West Point and instead chose to pursue a music career because of his disillusionment with his colleagues' values and because he knew West Point would not tolerate his dissidence.

Upon completing his term of enlistment, he studied art at Kendall College of Art and Design in Grand Rapids, Michigan. In 1988, he relocated to Los Angeles where his love of animals led him to practice interior design for pet stores, before he was quickly fired and began working in set construction. During the 1980s, Keenan played bass guitar for TexA.N.S. and sang for Children of the Anachronistic Dynasty, both independent bands.

After moving to Los Angeles, Keenan met Adam Jones who had heard him singing on a demo in college. Impressed with Keenan's vocals, Jones suggested that they form a band. Although reluctant at first, Keenan agreed and, in 1990, Tool was formed. Fronted by Keenan, the eventual lineup included guitarist Jones; his neighbor, drummer Danny Carey; and bassist Paul D'Amour, who would later be replaced by Justin Chancellor.

Tool signed to Zoo Entertainment in November 1991 and released the Opiate EP the following year. To support this release, the band toured with Fishbone and Rage Against the Machine. During a 1993 show at the Church of Scientology’s Celebrity Centre in Los Angeles, Keenan (who would later write the lyrics "Fuck L. Ron Hubbard and fuck all his clones" in "Ænema") bleated like a sheep for a good portion of the show. "Before our set this guy tries to intimidate me by showing me a gun in his jacket," Keenan recalled. "I explained to him that if he pissed me off I’d start a riot."

Shortly thereafter, Tool released their 1993 debut album, Undertow, in the United States. It was certified gold after just eight months, and platinum less than a year later. In 1994, the band released their single "Prison Sex"—a song in which Keenan opted to deliver in a slight grunge-style—with a corresponding music video created and directed by Jones. The video was deemed "too graphic and offensive", and was withdrawn by MTV after a few airings due to "a symbolic dealing with the sensitive subject of child abuse".

In October 1996, the band released their second studio album, Ænima. The album was certified gold in ten weeks, achieved double platinum in ten months, and won the Grammy Award for Best Metal Performance in 1998. After the release of the album, Tool began a prolonged legal battle with their label, Volcano Records (formerly Zoo Records), over contract violations. Following this legal battle, which resulted in a new three-record deal, the members of Tool decided to take some time off. During the hiatus, the band members were outspokenly critical of filesharing networks, due to the negative financial impact on artists dependent on success in record sales. During an interview with NY Rock in 2000, Keenan stated, "I think there are a lot of other industries out there that might deserve being destroyed. The ones who get hurt by MP3s are not so much companies or the business, but the artists, people who are trying to write songs."

Five years after the release of Ænima, Tool announced a new album, Systema Encéphale, with a 12-song tracklist in January 2001. A month later, the band revealed that the new album was actually titled Lateralus and that the previous announcement had been a ruse. The album was released in May 2001 to positive reviews. Known for his "dark, intelligent, compelling, and unexpected lyrical twists", Keenan was acclaimed for his songwriting on the album, in which he "doesn’t cross the line from darkness to ugliness ... as often as he has in the past". In an interview with NY Rock, Keenan explained, "Everything we release with Tool is inspired by our music. It doesn't matter if it is a video or if its lyrics. The lyrics for "Schism" are nothing more than my interpretation of the music." The album became a worldwide success, reaching #1 on the U.S. Billboard 200 albums chart in its debut week, and Tool received their second Grammy Award for the best metal performance of 2001 for "Schism". In 2002, Keenan recorded a song called "Fallen" with 30 Seconds to Mars that was released on the band's self-titled debut album.